FIGURE 2.

ASF/SF2 and SC35 bind to ESE2 in vitro. A, RNA affinity chromatography. RNA molecules were covalently linked to Sepharose beads, and SR proteins purified from calf thymus were passed over the column followed by extensive washing. Bound proteins were eluted with increasing salt concentrations (100, 150, 200, 250, 300, 400, 500, and 500 mm) and a final elution with 8 m urea. Elution fractions were run on SDS-PAGE gels and stained with Coomassie Blue. B, RNA affinity chromatography was performed using either wild-type ESE2 (5′-AAAGGAACAGAAGUA-3′) or mutant ESE2 (5′-AUAGUAAUAGUAGUA-3′) RNAs. Individual bands were cut out of the gel and identified by mass spectrometry as ASF/SF2 and SC35.